Comparing Two Saliva Collection Methods to Evaluate Transcriptomic Profile and Microbial Contamination
Saliva contains a spectrum of RNAs that are contained inside EVs (extracellular vesicles) and within RNA binding complexes. These RNAs have the potential to serve as useful biomarkers as we can examine thousands of RNAs at one time, in addition to modifications such as editing sites and splice variants during transcription. However, quality of the RNA in saliva samples bring challenges due to RNases and bacterial contamination. The goal of this experiment is to find a saliva collection method that yields higher RNA quality. RNA extraction from biofluids allows analysis of gene expression. The importance of this experiment is that the analysis of gene expression may be used to diagnose and reveal biological functions of diseases and traumas, such as a concussion. This experiment compares two saliva collection methods. The tube collection method involves subjects to expectorate (“spit”) an amount of 2 mL in a collection tube. The RNAPro⋅SAL™method utilizes a device where collection is from the pool of saliva in the oral cavity to harvest a higher yield of saliva for transcriptomics. After saliva collection, the RNA of each of the samples were stabilized and isolated using the RNEASY Protect Saliva kit by Qiagen. Subsequently, the samples were purified, quantified, and prepped to cDNA. Finally, the cDNA was quantified, sequenced, and analyzed. Analysis concluded that microbial contamination was present in both collection methods. In addition, the order of the collection method for each sample potentially correlates with bacterial contamination and is worth investigating in the future for applications for saliva diagnostics. It is worth noting that the tube collection method yields higher human transcriptomic diversity, especially in brain related information compared to the RNAPro⋅SAL™. Future work will look to determine which method may be more efficient in evaluating information relating to neurodegenerative diseases and brain related traumas
